We propose to study the physiological factors that control the physical properties of the inner membranes of liver and muscle mitochondria and their significance for metabolic regulation. We plan to expand our studies of change in pH and membrane potentials and their effects on oxidative phosphorylation and other mitochondrial functions. We want to investigate to what extent, and how, these parameters can be regulated by physiological factors such as Ca(II), pH key metabolites, temperature, hormones and pathological conditions. We shall study the mitochondrial membrane surface charge with fluorescent probes and microelectrophoresis, surface pH with fluorescent pH probes, membrane phase separations and phase transitions by differential scanning calorimetry and lipid fluidity by fluorescence polarization, lipid spin probes, 31P-NMR and other physical techniques. The proton permeability of the membrane, the efficiency of energy conversion and other paramters related to oxidative phosphorylation would be correlated with the physical properties of the mitochondrial membranes. We shall investigate the effect of these parameters on mitochondrial function, their relation to delta pH and delta psi and the extent to which these parameters are regulated by phsyiological factors. In particular, the effect of glucagon and insulin on the physical state of the membrane and the possible alteration due to diabetes would be investigated. The results of these studies might lead to a better understanding of the regulation of mitochondrial function both in normal and pathological conditions. We hope to find out whether local membrane parameters are physiologically regulated so as to control mitochondrial function and what is their relation to the energetic regulation by the proton electrochemical gradient. Moreover, these studies should also contribute to a better understanding of the basic mechanism underlying energy conversion in oxidative phosphorylation by mitochondria.